Iron depletion results in Src kinase inhibition with associated cell cycle arrest in neuroblastoma cells.

Iron is required for cellular proliferation. Recently, using systematic time studies of neuroblastoma cell growth, we better defined the G1 arrest caused by iron chelation to a point in mid-G1, where cyclin E protein is present, but the cyclin E/CDK2 complex kinase activity is inhibited. In this study, we again used the neuroblastoma SKNSH cells lines to pinpoint the mechanism responsible for this G1 block. Initial studies showed in the presence of DFO, these cells have high levels of p27 and after reversal of iron chelation p27 is degraded allowing for CDK2 kinase activity. The initial activation of CDK2 kinase allows cells to exit G1 and enter S phase. Furthermore, we found that inhibition of p27 degradation by DFO is directly associated with inhibition of Src kinase activity measured by lack of phosphorylation of Src at the 416 residue. Activation of Src kinase occurs very early after reversal from the DFO G1 block and is temporally associated with initiation of cellular proliferation associated with entry into S phase. For the first time therefore we show that iron chelation inhibits Src kinase activity and this activity is a requirement for cellular proliferation.

Two cell cycle blocks caused by iron chelation of neuroblastoma cells: separating cell cycle events associated with each block.

Studies have presented evidence that besides the well described S phase block, treatment of cancer cell lines with the iron chelator deferrioxamine (DFO) also results in an earlier block in G1 phase. In this article, measurements of cell cycle regulatory proteins define this block at a very specific point in G1. DFO treatment results in markedly decreased cyclin A protein levels. Cyclin E levels that accumulate in early to mid-G1 are increased in cells treated with DFO as compared to the resting cells. The DFO S phase block is shown after cells are arrested at G1/S by (aphidicolin) then released into DFO. The same S phase block occurs with DFO treatment of a neuroblastoma cell line relatively resistant to the G1 DFO block. These experiments clearly differentiate the S phase DFO block from the earlier block pinpointed to a point in mid-G1, before G1/S when cyclin E protein increases but before increased cyclin A synthesis. Apoptosis was observed in cells inhibited by DFO at both cell cycle arrest points.

Single-dosage pharmacokinetics of sodium ferric gluconate complex in iron-deficient pediatric hemodialysis patients.

BACKGROUND AND OBJECTIVES: The clinical use of sodium ferric gluconate complex in iron-deficient pediatric patients receiving hemodialysis was recently approved. This study was designed to describe the pharmacokinetic parameters of the medication. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Iron-deficient pediatric (< or = 15 yr) hemodialysis patients were randomly assigned to two doses (1.5 and 3.0 mg/kg) of sodium ferric gluconate complex. Blood samples taken during a 1-h infusion and at multiple intervals during 48 h were analyzed for total iron, transferrin-bound iron, and sodium ferric gluconate complex-bound iron. RESULTS: Forty-nine patients (mean age 12.3 +/- 2.5 yr) participated in the study. Mean serum iron concentrations rapidly increased in a dosage-dependent manner. A rapid rise in total serum iron was followed by a slower, less prominent rise in transferrin-bound iron. This was qualitatively confirmed by visualization of the transferrin bands from polyacrylamide gel electrophoresis. Single-dose pharmacokinetics of sodium ferric gluconate complex-bound iron was described using noncompartmental analytical methods. Mean values for the 1.5 mg/Kg dose were as follows: t(1/2) 2.0 +/- 0.7 h, Cmax 1287 mcg/dl, Tmax 1.1 +/- 0.23 h, Cl 0.69 +/- 0.50 L/h, Vd 1.6 +/- 0.6 L, AUC(0-infinity). 9499 +/- 4089 mcg x hr/dl. CONCLUSIONS: The infusion of sodium ferric gluconate complex to pediatric patients who receive hemodialysis appears to result in a delayed transfer of iron to transferrin, likely after an initial movement through the reticuloendothelial system. Differences noted between the pediatric and adult pharmacokinetic data may result from the unique aspects of the study populations and the respective study designs.

Expression of vascular endothelial growth factor and proliferation marker MIB1 are influenced by neoadjuvant chemotherapy in locally advanced breast cancer.

Neoadjuvant chemotherapy (NACT) has become the standard of care for patients with locally advanced breast cancer (LABC). This was a retrospective review of 21 consecutive women who received NACT as initial treatment of LABC, followed by surgical excision. The pre- and post-treatment breast specimens and post-treatment axillary lymph nodes with metastases were immunostained to evaluate for proliferative index (PI) (MIB-1 Immunotech) and vascular endothelial growth factor (VEGF) expression (Santa Cruz, CA, clone A-20). Thirteen of the 21 patients (62%) had more than 50% tumor shrinkage following NACT. The breast's mean PI decreased from 47.86% to 23.95% after treatment (P = 0.005). The mean PI in the post-treatment lymph nodes was 24.47%. A nodal post-NACT PI of less than 10% and progesterone receptor-positive tumor status were associated with better survival, as all such patients are alive. A high PI after NACT was associated with recurrence or death. All of the patients who showed an excellent clinical response had either a decrease in the PI or an absence of a high level of VEGF after NACT. Most patients exhibited persistent expression of VEGF after NACT. Pathologic response in the primary tumor did not correlate with the response in the lymph nodes or with overall survival. NACT reduces the size and PI of the primary breast tumor independent of the patient's node status. The PI may be an early means by which to identify tumors most likely to reduce in size with chemotherapy. A low PI after NACT is associated with better survival. There is persistent expression of VEGF in post-NACT residual breast carcinoma. Thus, anti-VEGF drugs after conventional chemotherapy may benefit patients with residual carcinoma.

Single-dose pharmacokinetics of sodium ferric gluconate complex in iron-deficient subjects.

STUDY OBJECTIVES: To determine the single-dose pharmacokinetics of intravenous sodium ferric gluconate complex in sucrose injection (SFGC) in iron-deficient human volunteers, and to assess iron transport. DESIGN: Open-label, randomized study. SETTING: Clinical research facility. SUBJECTS: Fourteen iron-deficient men and women. INTERVENTIONS: Subjects were randomized to receive a single intravenous dose of either SFGC 62.5 mg administered over 30 minutes or SFGC 125 mg over 60 minutes. Five days later, the same subjects were rerandomized to receive a second intravenous dose of SFGC, either 62.5 mg administered over 4 minutes or 125 mg over 7 minutes. MEASUREMENTS AND MAIN RESULTS: Blood samples were collected at predefined times before, during, and up to 72 hours after the infusion to determine the single-dose pharmacokinetics of SFGC. Assays were performed for both total iron and transferrin-bound iron, from which drug-bound iron could be calculated. Urine was collected over 24 hours before dosing and for 24 hours after the start of infusion to determine the renal elimination of iron. Clearance of SFGC from serum was rapid and far exceeded rates reported for iron dextran. Pharmacokinetic parameters were unaffected by dose or infusion rate. Serum iron derived from SFGC did not exceed the binding capacity of transferrin. Serum iron from SFGC became rapidly available (< 24 hrs) as transferrin-bound iron, but only after passage through another compartment, presumably the reticuloendothelial system (RES). At least 80% of the administered iron was transported to bone marrow within 24 hours after infusion. CONCLUSIONS: Iron derived from SFGC appears to be rapidly transferred to a bioavailable iron compartment as transferrin-bound iron after digestion in the RES. At the doses administered in this study, liberation of potentially toxic, free iron was not detectable.

Clinical evaluation of heme iron polypeptide: sustaining a response to rHuEPO in hemodialysis patients.

BACKGROUND: Optimizing iron and recombinant human erythropoietin (rHuEPO) therapy is necessary to achieve target hemoglobin levels and minimize costs as the end-stage renal disease (ESRD) population expands. Oral iron products in patients with ESRD have been largely abandoned, and the safety of intravenous (IV) iron preparations has improved with the introduction of new-generation compounds that have little allergenicity. Recent work suggests oral heme iron may be an effective supplement for hemodialysis (HD) patients because it is absorbed by patients with high ferritin levels, has fewer side effects, and its absorption is stimulated by erythropoietin administration. METHODS: We performed an open, 6-month, prospective evaluation of heme iron in HD patients who had been on maintenance IV iron therapy. IV iron was discontinued and replaced with oral heme iron. Serum iron level, hematocrit (Hct), and erythropoietin and IV iron dose were monitored. RESULTS: During 6 months, 4 of 37 patients (11%) dropped out because of insufficient iron supplementation or intolerance and 5 patients (14%) were dropped because of unrelated complications or protocol violation. A slight reduction in average transferrin saturation (TSAT) was seen early, but reversed, and no significant changes were seen in TSAT or Hct. A significant reduction in average serum ferritin level was seen at months 4 through 6 (P < 0.01). CONCLUSION: During the 6-month study period, heme iron polypeptide successfully replaced IV iron therapy in a majority of HD patients and maintained target Hcts with no concomitant use of IV iron. This treatment was associated with a significant increase in rHuEPO efficiency (P = 0.04).